Sunlight has a significant effect on the skin causing premature aging, skin cancer and a host of other skin changes such as erythema and tanning. The majority of the damage caused by sunlight is attributed to ultraviolet (UV) radiation, which has a wavelength from 200 nm to 400 nm. Ultraviolet radiation is divided into three categories, UVA, UVB or UVC, depending on wavelength. UVA, which has a wavelength range from 320-400 nm, can cause tanning and mild sunburn. UVB, which has a wavelength range from 290-320 nm, can cause sunburn and stimulate pigmentation. UVC, which has a wavelength range from 100-290 nm, can cause damage but not tanning. Exposure of the skin to UV radiation induces biphasic reactions. Thus, upon initial exposure an immediate erythema reaction occurs, which is a weak reaction that fades within 30 minutes. A delayed erythema reaction occurs after 2-5 hours of exposure and peaks around 10-24 hours. Enhanced prostaglandin and leukotriene production are the major mechanisms of action for UV, sun and chemical/thermal caused erythema. (Wang (2002) Adv. Dermatol. 18:247).
The liberation and metabolism of arachidonic acid (AA) from the cell membrane results in the generation of pro-inflammatory metabolites by several different pathways. Arguably, two of the most important pathways to inflammation are mediated by the enzymes lipoxygenase (LOX) and cyclooxygenase (COX). These are parallel pathways that result in the generation of leukotrienes and prostaglandins, respectively, which play important roles in the initiation and progression of the inflammatory response. These vasoactive compounds are chemotaxins, which both promote infiltration of inflammatory cells into tissues and serve to prolong the inflammatory response. Consequently, the enzymes responsible for generating these mediators of inflammation have become the targets in the current invention to develop topically administered therapeutic agents aimed at the dual inhibition of inflammation resulting from both pathways which contribute to the physiological and pathological processes of diseases and conditions such as sun burn, thermal burns, scald, acne, topical wounds, lupus erythromatosus, psoriasis, carcinoma, melanoma, and other mammalian skin cancers.
Inhibition of the COX enzyme is the mechanism of action attributed to most nonsteroidal anti-inflammatory drugs (NSAIDS). There are two distinct isoforms of the COX enzyme (COX-1 and COX-2), which share approximately 60% sequence homology, but differ in expression profiles and function. COX-1 is a constitutive form of the enzyme that has been linked to the production of physiologically important prostaglandins, which help regulate normal physiological functions, such as platelet aggregation, protection of cell function in the stomach and maintenance of normal kidney function. (Dannhardt and Kiefer (2001) Eur. J. Med. Chem. 36:109-26). The second isoform, COX-2, is a form of the enzyme that is inducible by pro-inflammatory cytokines, such as interleukin-1β (IL-1β) and other growth factors. (Herschmann (1994) Cancer Metastasis Rev. 134:241-56; Xie et al. (1992) Drugs Dev. Res. 25:249-65). This isoform catalyzes the production of prostaglandin E2 (PGE2) from arachidonic acid (AA). Inhibition of COX is responsible for the anti-inflammatory activity of conventional NSAIDs.
Inhibitors that demonstrate dual specificity for COX and LOX would have the obvious benefit of inhibiting multiple pathways of arachidonic acid metabolism. Such inhibitors would block the inflammatory effects of prostaglandins (PG), as well as, those of multiple leukotrienes (LT) by limiting their production. This includes the vasodilation, vasopermeability and chemotactic effects of PGE2, LTB4, LTD4 and LTE4, also known as the slow reacting substance of anaphalaxis. Of these, LTB4 has the most potent chemotactic and chemokinetic effects. (Moore (1985) in Prostanoids: pharmacological, physiological and clinical relevance, Cambridge University Press, N.Y., pp. 229-230).
Because the mechanism of action of COX inhibitors overlaps that of most conventional NSAID's, COX inhibitors are used to treat many of the same symptoms, including pain and swelling associated with inflammation in transient conditions and chronic diseases in which inflammation plays a critical role. Transient conditions include treatment of inflammation associated with minor abrasions or contact dermatitis, as well as, skin conditions that are directly associated with the prostaglandin and leukotriene pathways, such as skin hyperpigmentation, age spots, vitilago, systemic lupus erythromatosus, psoriasis, carcinoma, melanoma, and other mammalian skin cancers. The use of COX inhibitors has been expanded to include diseases, such as systemic lupus erythromatosus (SLE) (Goebel et al. (1999) Chem. Res. Toxicol. 12:488-500; Patrono et al. (1985) J. Clin. Invest. 76:1011-1018), as well as, rheumatic skin conditions, such as scleroderma. COX inhibitors are also used for the relief of inflammatory skin conditions that are not of rheumatic origin, such as psoriasis, in which reducing the inflammation resulting from the overproduction of prostaglandins could provide a direct benefit. (Fogh et al. (1993) Acta Derm Venerologica 73:191-193). Recently over expression of 5-lipoxygenase in the skin of patients with system sclerosis has been reported. This has led to the suggestion that the LOX pathway may be of significance in the pathogenesis of system sclerosis and may represent a valid therapeutic target. (Kowal-Bielecka (2001) Arthritis Rheum. 44(8):1865). Finally, the increased enzymatic activity of both the COX-2 and 5-LOX at the site of allergen injections suggests the potential for using dual COX/LOX inhibitors to treat the symptoms of both the early and late phases of the skin allergic response. (Church (2002) Clin. Exp. Allergy. 32(7):1013).
Topical application of a selective cyclooxygenase inhibitor has been shown to suppress UVB mediated cutaneous inflammation following both acute and long-term exposure. Additionally, edema, dermal neutrophil infiltration and activation, PGE2 levels and the formation of sunburn cells were reduced by the topical application of a COX inhibitor. (Wilgus (2000) Prostaglandins Other Lipid Mediat. 62(4):367). The COX inhibitor Celebrex™ has been shown to reduce the effects of UV induced inflammation when administered systematically (Wilgus et al. (2002) Adv. Exp. Med. Biol. 507:85), and topically (Wilgus et al. (2000) Protaglandins Other Lipid Mediat. 62:367). In animal models, the known COX inhibitor aspirin and various lipoxygenase inhibitors exhibited vasoprotective activity against inflammation and vasodepression resulting from UV irradiation. (Kuhn (1988) Biomed. Biochim. Acta. 47:S320). Acute or long-term chronic UV exposure causes skin damage and photoageing that are characterized by degradation of collagen and accumulation of abnormal elastin in the superficial dermis. A dual COX/LOX inhibitor can be utilized to prevent and treat collagen degradation caused by inflammatory infiltration by significantly reducing the vasodilating, vasopermeability, chemotactic and chemotaxins-prostaglandins (PG), as well as, those of multiple leukotrienes (LT). (Bosset (2003) Br. J. Dermatol. 149(4):826; Hase (2000) Br. J. Dermatol. 142(2):267). Additionally, chemically induced oxidative stress in mouth skin can be inhibited by separately administrating COX and LOX inhibitors to reduce leukocyte adhesion, infiltration and H2O2 generation. (Nakamura (2003) Free Radical Biol. Med. 35(9):997).
In addition to their use as anti-inflammatory agents, another potential role for COX inhibitors is in the treatment of cancer. Over expression of COX has been demonstrated in various human malignancies and inhibitors of COX have been shown to be efficacious in the treatment of animals with skin tumors. While the mechanism of action is not completely understood, the over expression of COX has been shown to inhibit apoptosis and increase the invasiveness of tumorgenic cell types. (Dempke et al. (2001) J. Can. Res. Clin. Oncol. 127:411-17; Moore and Simmons (2000) Current Med. Chem. 7:1131-1144). Up regulated COX production has been implicated in the generation of actinic keratosis and squamous cell carcinoma in skin. Increased amounts of COX were also found in lesions produced by DNA damage. (Buckman et al. (1998) Carcinogenesis 19:723). Therefore, control of expression or protein function of COX would seem to lead to a decrease in the inflammatory response and the eventual progression to cancer. In fact, COX inhibitors such as indomethacin and Celebrex™ have been found to be effective in treating UV induced erythema and tumor formation. (Fischer (1999) Mol. Carcinog. 25:231; Pentland (1999) Carcinogenesis 20:1939). Recently, the over expression of lipoxygenase has also been shown to be related to epidermal tumor development (Muller (2002) Cancer Res. 62(16):4610) and melanoma carcinogenesis (Winer (2002) Melanoma Res. 12(5):429). The arachidonic acid (AA) metabolites generated from lipoxygenase pathways play important roles in tumor growth related signal transduction suggesting that that the inhibition of lipoxygenase pathways should be a valid target to prevent cancer progression. (Cuendet (2000) Drug Metabol Drug Interact 17(4):109; Steele (2003) Mutat Res. 523-524:137). Thus, the use of therapeutic agents having dual COX/LOX inhibitory activity offers significant advantages in the chemoprevention of cancer.
Prostaglandins and leukotrienes also play important roles in the physiological and pathological processes of wounds, burns, scald, acne, microbial infections, dermatitis, and many other diseases and conditions of the skin. The activation of a pro-inflammatory cascade after thermal or chemical burns with significantly elevated cyclooxygenase and lipoxygenase activities are well documented and play an important role in the development of subsequent severe symptoms and immune dysfunction that may lead to multiple organ failure. (Schwacha (2003) Burns 29(1):1; He (2001) J. Burn Care Rehabil. 22(1):58).
Acne is a disease of the pilosebaceous unit with abnormalities in sebum production, follicular epithelial desquamation, bacterial proliferation and inflammation. The inflammatory properties of acne can be detected by polarized light photography and utilized for clinical diagnosis, including an evaluation of the extent of the acne and also to determine the effectiveness of therapy. (Phillips (1997) J. Am. Acad. Dermatol. 37(6):948). Current therapeutic agents for the prevention and treatment of acne include anti-inflammatory agents, like retinoids, antimicrobial agents and hormonal drugs. (Leyden (2003) J. Am. Acad. Dermatol. 49(3 Suppl):5200). Topical application of anti-inflammatory drugs, such as retinoids (Millikan (2003) J. Am. Acad. Dermatol. 4(2):75) and the COX inhibitor salicylic acid (Lee (2003) Dermatol Surg 29(12):1196) have been clinically demonstrated as an effective and safe therapy for the treatment of acne. Additionally, the use of nonsteroidal anti-inflammatory drugs (NSAIDs) are well documented as therapeutic agents for common and uncommon dermatoses, including acne, psoriasis, sun burn, erythema nodosum, cryoglobulinemia, Sweet's syndrome, systemic mastocytosis, urticarial, liverdoid and nodular vasculitis. (Friedman (2002) J. Cutan Med. Surg. 6(5):449).
Flavonoids or bioflavonoids are a widely distributed group of natural products, which have been reported to have antibacterial, anti-inflammatory, antiallergic, antimutagenic, antiviral, antineoplastic, anti-thrombic and vasodilatory activity. The structural unit common to this group of compounds includes two benzene rings on either side of a 3-carbon ring as illustrated by the following general structural formula:
Various combinations of hydroxyl groups, sugars, oxygen and methyl groups attached to this general three ring structure create the various classes of flavonoids, which include flavanols, flavones, flavan-3-ols (catechins), anthocyanins and isoflavones.
Free-B-Ring flavones and flavonols are a specific class of flavonoids, which have no substituent groups on the aromatic B ring (referred to herein as Free-B-Ring flavonoids), as illustrated by the following general structure:
                wherein        R1, R2, R3, R4, and R5 are independently selected from the group consisting of —H, —OH, —SH, OR, —SR, —NH2, —NHR, —NR2, —NR3+X−, a carbon, oxygen, nitrogen or sulfur, glycoside of a single or a combination of multiple sugars including, but not limited to aldopentoses, methyl-aldopentose, aldohexoses, ketohexose and their chemical derivatives thereof;        wherein        R is an alkyl group having between 1-10 carbon atoms; and        X is selected from the group of pharmaceutically acceptable counter anions including, but not limited to hydroxyl, chloride, iodide, fluoride, sulfate, phosphate, acetate, carbonate, etc.        
Free-B-Ring flavonoids are relatively rare. Out of 9,396 flavonoids synthesized or isolated from natural sources, only 231 Free-B-Ring flavonoids are known (The Combined Chemical Dictionary, Chapman & Hall/CRC, Version 5:1 Jun. 2001). Free-B-Ring flavonoids have been reported to have diverse biological activity. For example, galangin (3,5,7-trihydroxyflavone) acts as antioxidant and free radical scavenger and is believed to be a promising candidate for anti-genotoxicity and cancer chemoprevention. (Heo et al. (2001) Mutat. Res. 488(2):135-150). It is an inhibitor of tyrosinase monophenolase (Kubo et al. (2000) Bioorg. Med. Chem. 8(7):1749-1755), an inhibitor of rabbit heart carbonyl reductase (Imamura et al. (2000) J. Biochem. 127(4):653-658), has antimicrobial activity (Afolayan and Meyer (1997) Ethnopharmacol. 57(3):177-181) and antiviral activity (Meyer et al. (1997) J. Ethnopharmacol. 56(2):165-169). Baicalein and two other Free-B-Ring flavonoids, have antiproliferative activity against human breast cancer cells. (So et al. (1997) Cancer Lett. 112(2):127-133).
Typically, flavonoids have been tested for biological activity randomly based upon their availability. Occasionally, the requirement of substitution on the B-ring has been emphasized for specific biological activity, such as the B-ring substitution required for high affinity binding to p-glycoprotein (Boumendjel et al. (2001) Bioorg. Med. Chem. Lett. 11(1):75-77); cardiotonic effect (Itoigawa et al. (1999) J. Ethnopharmacol. 65(3): 267-272), protective effect on endothelial cells against linoleic acid hydroperoxide-induced toxicity (Kaneko and Baba (1999) Biosci Biotechnol. Biochem 63(2):323-328), COX-1 inhibitory activity (Wang (2000) Phytomedicine 7:15-19) and prostaglandin endoperoxide synthase (Kalkbrenner et al. (1992) Pharmacology 44(1):1-12). Only a few publications have mentioned the significance of the unsubstituted B ring of the Free-B-Ring flavonoids. One example, is the use of 2-phenyl flavones, which inhibit NADPH quinone acceptor oxidoreductase, as potential anticoagulants. (Chen et al. (2001) Biochem. Pharmacol. 61(11):1417-1427).
The mechanism of action with respect to the anti-inflammatory activity of various Free-B-Ring flavonoids has been controversial. The anti-inflammatory activity of the Free-B-Ring flavonoids, chrysin (Liang et al. (2001) FEBS Lett. 496(1):12-18), wogonin (Chi et al. (2001) Biochem. Pharmacol. 61:1195-1203) and halangin (Raso et al. (2001) Life Sci. 68(8):921-931), has been associated with the suppression of inducible cyclooxygenase and nitric oxide synthase via activation of peroxisome proliferator activated receptor gamma (PPARγ) and influence on degranulation and AA release. (Tordera et al. (1994) Z. Naturforsch [C] 49:235-240). It has been reported that oroxylin, baicalein and wogonin inhibit 12-lipoxygenase activity without affecting cyclooxygenase. (You et al. (1999) Arch. Pharm. Res. 22(1):18-24). More recently, the anti-inflammatory activity of wogonin, baicalin and baicalein has been reported as occurring through inhibition of inducible nitric oxide synthase and cox-2 gene expression induced by nitric oxide inhibitors and lipopolysaccharide. (Chen et al. (2001) Biochem. Pharmacol. 61(11):1417-1427). It has also been reported that oroxylin acts via suppression of NFκB activation. (Chen et al. (2001) Biochem. Pharmacol. 61(11):1417-1427). Finally, wogonin reportedly inhibits inducible PGE2 production in macrophages. (Wakabayashi and Yasui (2000) Eur. J. Pharmacol. 406(3):477-481).
Inhibition of the phosphorylation of mitrogen-activated protein kinase and inhibition of Ca2+ ionophore A23187 induced PGE2 release by baicalein has been reported as the mechanism of anti-inflammatory activity of Scutellariae radix. (Nakahata et al. (1999) Nippon Yakurigaku Zasshi, 114, Supp. 11:215P-219P; Nakahata et al. (1998) Am. J. Chin Med. 26:311-323). Baicalin from Scutellaria baicalensis, reportedly inhibits superantigenic staphylococcal exotoxins stimulated T-cell proliferation and production of IL-1β, IL-6, tumor necrosis factor-α (TNF-α), and interferon-γ (IFN-γ). (Krakauer et al. (2001) FEBS Lett. 500:52-55). Thus, the anti-inflammatory activity of baicalin has been associated with inhibiting the pro-inflammatory cytokines mediated signaling pathways activated by superantigens. However, it has also been proposed that the anti-inflammatory activity of baicalin is due to the binding of a variety of chemokines, which limits their biological activity. (Li et al. (2000) Immunopharmacology 49:295-306). Recently, the effects of baicalin on adhesion molecule expression induced by thrombin and thrombin receptor agonist peptide (Kimura et al. (2001) Planta Med. 67:331-334), as well as, the inhibition of mitogen-activated protein kinase cascade (MAPK) (Nakahata et al. (1999) Nippon Yakurigaku Zasshi, 114, Supp 11:215P-219P; Nakahata et al. (1998) Am. J. Chin Med. 26:311-323) have been reported.
The Chinese medicinal plant, Scutellaria baicalensis contains significant amounts of Free-B-Ring flavonoids, including baicalein, baicalin, wogonin and baicalenoside. Traditionally, this plant has been used to treat a number of conditions including clearing away heat, purging fire, dampness-warm and summer fever syndromes; polydipsia resulting from high fever; carbuncle, sores and other pyogenic skin infections; upper respiratory infections, such as acute tonsillitis, laryngopharyngitis and scarlet fever; viral hepatitis; nephritis; pelvitis; dysentery; hematemesis and epistaxis. This plant has also traditionally been used to prevent miscarriage. (Encyclopedia of Chinese Traditional Medicine, ShangHai Science and Technology Press, ShangHai, China, 1998). Clinically Scutellaria is now used to treat conditions such as pediatric pneumonia, pediatric bacterial diarrhea, viral hepatitis, acute gallbladder inflammation, hypertension, topical acute inflammation, resulting from cuts and surgery, bronchial asthma and upper respiratory infections. (Encyclopedia of Chinese Traditional Medicine, ShangHai Science and Technology Press, ShangHai, China, 1998). The pharmacological efficacy of Scutellaria roots for treating bronchial asthma is reportedly related to the presence of Free-B-Ring flavonoids and their suppression of eotaxin associated recruitment of eosinophils. (Nakajima et al. (2001) Planta Med. 67(2):132-135).
To date, a number of naturally occurring Free-B-Ring flavonoids have been commercialized for varying uses. For example, liposome formulations of Scutellaria extracts have been utilized for skin care (U.S. Pat. Nos. 5,643,598; 5,443,983). Baicalin has been used for preventing cancer, due to its inhibitory effects on oncogenes (U.S. Pat. No. 6,290,995). Baicalin and other compounds have been used as antiviral, antibacterial and immunomodulating agents (U.S. Pat. No. 6,083,921 and WO98/42363) and as natural anti-oxidants (WO98/49256 and Poland Pub. No. 9,849,256). Scutellaria baicalensis root extract has been formulated as a supplemental sun screen agent with additive effects of the cumulative SPFs of each individual component in a topical formulation (WO98/19651). Chrysin has been used for its anxiety reducing properties (U.S. Pat. No. 5,756,538). Anti-inflammatory flavonoids are used for the control and treatment of anorectal and colonic diseases (U.S. Pat. No. 5,858,371), and inhibition of lipoxygenase (U.S. Pat. No. 6,217,875). These compounds are also formulated with glucosamine collagen and other ingredients for repair and maintenance of connective tissue (U.S. Pat. No. 6,333,304). Flavonoid esters constitute active ingredients for cosmetic compositions (U.S. Pat. No. 6,235,294). U.S. application Ser. No. 10/091,362, filed Mar. 1, 2002, entitled “Identification of Free-B-Ring Flavonoids as Potent COX-2 Inhibitors,” and U.S. application Ser. No. 10/427,746, filed Jul. 22, 2003, entitled “Formulation of a Mixture of Free-B-Ring Flavonoids and Flavans as a Therapeutic Agent” both disclose a method for inhibiting the cyclooxygenase enzyme COX-2 by administering a composition comprising a Free-B-Ring flavonoid or a composition containing a mixture of Free-B-Ring flavonoids to a host in need thereof. This is the first report of a link between Free-B-Ring flavonoids and COX-2 inhibitory activity. These applications are specifically incorporated herein by reference in their entirety.
Japanese Pat. No. 63027435, describes the extraction, and enrichment of baicalein and Japanese Pat. No. 61050921 describes the purification of baicalin.
Flavans include compounds illustrated by the following general structure:
                wherein        R1, R2, R3, R4 and R5 are independently selected from the group consisting of —H, —OH, —SH, —OCH3, —SCH3, —OR, —SR, —NH2, —NRH, —NR2, —NR3+X−, esters of the mentioned substitution groups, including, but not limited to, gallate, acetate, cinnamoyl and hydroxyl-cinnamoyl esters, trihydroxybenzoyl esters and caffeoyl esters, and their chemical derivatives thereof; a carbon, oxygen, nitrogen or sulfur glycoside of a single or a combination of multiple sugars including, but not limited to, aldopentoses, methyl aldopentose, aldohexoses, ketohexose and their chemical derivatives thereof; dimer, trimer and other polymerized flavans;        wherein        R is an alkyl group having between 1-10 carbon atoms; and        X is selected from the group of pharmaceutically acceptable counter anions including, but not limited to hydroxyl, chloride, iodide, sulfate, phosphate, acetate, fluoride, and carbonate, etc.        
Catechin is a flavan, found primarily in green tea, having the following structure:
Catechin works both alone and in conjunction with other flavonoids found in tea, and has both antiviral and antioxidant activity. Catechin has been shown to be effective in the treatment of viral hepatitis. It also appears to prevent oxidative damage to the heart, kidney, lungs and spleen and has been shown to inhibit the growth of stomach cancer cells.
Catechin and its isomer epicatechin inhibit prostaglandin endoperoxide synthase with an IC50 value of 40 μM. (Kalkbrenner et al. (1992) Pharmacol. 44:1-12). Five flavan-3-ol derivatives, including (+)-catechin and gallocatechin, isolated from four plant species: Atuna racemosa, Syzygium carynocarpum, Syzygium malaccense and Vantanea peruviana, exhibit equal to weaker inhibitory activity against COX-2, relative to COX-1, with IC50 values ranging from 3.3 μM to 138 μM (Noreen et al. (1998) Planta Med. 64:520-524). (+)-Catechin, isolated from the bark of Ceiba pentandra, inhibits COX-1 with an IC50 value of 80 μM (Noreen et al. (1998) J. Nat. Prod. 61:8-12). Commercially available pure (+)-catechin inhibits COX-1 with an IC50 value of around 183 to 279 μM depending upon the experimental conditions, with no selectivity for COX-2. (Noreen et al. (1998) J. Nat. Prod. 61:1-7).
Green tea catechin, when supplemented into the diets of Sprague dawley male rats, lowered the activity level of platelet PLA2 and significantly reduced platelet cyclooxygenase levels. (Yang et al. (1999) J. Nutr. Sci. Vitaminol. 45:337-346). Catechin and epicatechin reportedly weakly suppress cox-2 gene transcription in human colon cancer DLD-1 cells (IC50=415.3 μM). (Mutoh et al. (2000) Jpn. J. Cancer Res. 91:686-691). The neuroprotective ability of (+)-catechin from red wine results from the antioxidant properties of catechin, rather than inhibitory effects on intracellular enzymes, such as cyclooxygenase, lipoxygenase, or nitric oxide synthase (Bastianetto et al. (2000) Br. J. Pharmacol. 131:711-720). Catechin derivatives purified from green and black tea, such as epigallocatechin-3-gallate (EGCG), epigallocatechin (EGC), epicatechin-3-gallate (ECG), and theaflavins showed inhibition of cyclooxygenase and lipoxygenase dependent metabolism of AA in human colon mucosa and colon tumor tissues (Hong et al. (2001) Biochem. Pharmacol. 62:1175-1183) and induce cox-2 expression and PGE2 production (Park et al. (2001) Biochem. Biophys. Res. Commun. 286:721-725). Epiafzelechin isolated from the aerial parts of Celastrus orbiculatus exhibited dose-dependent inhibition of COX-1 activity with an IC50 value of 15 μM and also demonstrated anti-inflammatory activity against carrageenin-induced mouse paw edema following oral administration at a dosage of 100 mg/kg. (Min et al. (1999) Planta Med. 65:460-462).
Acacia is a genus of leguminous trees and shrubs. The genus Acacia includes more than 1000 species belonging to the family of Leguminosae and the subfamily of Mimosoideae. Acacias are distributed worldwide in tropical and subtropical areas of Central and South America, Africa, parts of Asia, as well as, Australia, which has the largest number of endemic species. Acacias are very important economically, providing a source of tannins, gums, timber, fuel and fodder. Tannins, which are isolated primarily from bark, are used extensively for tanning hides and skins. Some Acacia barks are also used for flavoring local spirits. Some indigenous species like A. sinuata also yield saponins, which are any of various plant glucosides that form soapy lathers when mixed and agitated with water. Saponins are used in detergents, foaming agents and emulsifiers. The flowers of some Acacia species are fragrant and used to make perfume. The heartwood of many Acacias is used for making agricultural implements and also provides a source of firewood. Acacia gums find extensive use in medicine and confectionary and as sizing and finishing materials in the textile industry.
To date, approximately 330 compounds have been isolated from various Acacia species. Flavonoids are the major class of compounds isolated from Acacias. Approximately 180 different flavonoids have been identified, 111 of which are flavans. Terpenoids are second largest class of compounds isolated from species of the Acacia genus, with 48 compounds having been identified. Other classes of compounds isolated from Acacia include, alkaloids (28), amino acids/peptides (20), tannins (16), carbohydrates (15), oxygen heterocycles (15) and aliphatic compounds (10). (Buckingham, The Combined Chemical Dictionary, Chapman & Hall CRC, version 5:2, December 2001).
Phenolic compounds, particularly flavans are found in moderate to high concentrations in all Acacia species. (Abdulrazak et al. (2000) Journal of Animal Sciences. 13:935-940). Historically, most of the plants and extracts of the Acacia genus have been utilized as astringents to treat gastrointestinal disorders, diarrhea, indigestion and to stop bleeding. (Vautrin (1996) Universite Bourgogne (France) European abstract 58-01C:177; Saleem et al. (1998) Hamdard Midicus. 41:63-67). The bark and pods of Acacia arabica Willd. contain large quantities of tannins and have been utilized as astringents and expectorants. (Nadkarni (1996) India Materia Medica, Bombay Popular Prakashan, pp. 9-17). Diarylpropanol derivatives, isolated from stem bark of Acacia tortilis from Somalia, have been reported to have smooth muscle relaxing effects. (Hagos et al. (1987) Planta Medica. 53:27-31, 1987). It has also been reported that terpenoid saponins isolated from Acacia victoriae have an inhibitory effect on dimethylbenz(a)anthracene-induced murine skin carcinogenesis (Hanausek et al. (2000) Proceedings American Association for Cancer Research Annual Meeting 41:663) and induce apotosis (Haridas et al. (2000) Proceedings American Association for Cancer Research Annual Meeting. 41:600). Plant extracts from Acacia nilotica have been reported to have spasmogenic, vasoconstrictor and anti-hypertensive activity (Amos et al. (1999) Phytotherapy Research 13:683-685; Gilani et al. (1999) Phytotherapy Research. 13:665-669), and antiplatelet aggregatory activity (Shah et al. (1997) General Pharmacology. 29:251-255). Anti-inflammatory activity has been reported for A. nilotica. It was speculated that flavonoids, polysaccharides and organic acids were potential active components. (Dafallah and Al-Mustafa (1996) American Journal of Chinese Medicine. 24:263-269). To date, the only reported 5-lipoxygenase inhibitor isolated from Acacia is a monoterpenoidal carboxamide (Seikine et al. (1997) Chemical and Pharmaceutical Bulletin. 45:148-11).
The extract from the bark of Acacia has been patented in Japan for external use as a whitening agent (Abe, JP10025238), as a glucosyl transferase inhibitor for dental applications (Abe, JP07242555), as a protein synthesis inhibitor (Fukai, JP 07165598), as an active oxygen scavenging agent for external skin preparations (Honda, JP 07017847, Bindra U.S. Pat. No. 6,126,950), and as a hyaluronidase inhibitor for oral consumption to prevent inflammation, pollinosis and cough (Ogura, JP 07010768).
To date, Applicant is unaware of any reports of a formulation combining only Free-B-Ring-Flavonoids and flavans as the primary biologically active components for the dual inhibition of the COX/LOX enzymes that yield significant benefit to mammal skin conditions.